低含量血清培养IBRS-2细胞及其增殖猪细小病毒N株的研究

摘要

In this study, the MEM containing 8%, 4%, 2% and 1% sera, respectively, were used to culture IBRS-2 cells and to proliferate Porcine parvovirus(PPV) N strain. The results showed that IBRS-2 cells were in good growth state in concentrations of 8%, 4% and 2% sera culture medium, respectively, but when the concentration of the serum was decreased to 1%, the cells were in poor attachment and growth. Meanwhile, the densities of IBRS-2 cells at 0, 24, 48 ,72 and 96 h time point in concentrations of 8%, 4% and 2% serum culture medium was not significantly different with the control. Thereby, the culture media containing 2% serum were selected for cell cultivation and virus yield, and the virus titers were determined by TCID50and hemagglutination test. The results showed that the PPV N strain average titer of PPV N strain in 2% serum was not significantly different from that in routine culture (6%～10% serum) , indicating that the system could be used for the proliferation of PPV N strain. Results show that this study successfully domesticated out the IBRS-2 cells that could grow well under low serum culture conditions (containing 2% serum), and the PPV N strains could proliferate well in the lower serum cultured IBRS - 2 cells.%依次采用含8%、4%、2%、1%小牛血清MEM培养基驯化IBRS-2细胞,并在驯化好的IBRS-2细胞上接种猪细小病毒(Porcine parvovirus,PPV)N株,确定PPV N株在该培养体系下的生长情况.结果显示,用含8%、4%、2%小牛血清MEM培养基各进行5代次驯化,IBRS-2细胞能完全适应且生长状态良好.当血清浓度降至1%时,仅传代1次,细胞就无法保持良好状态,细胞出现贴壁较差、拉网、生长停滞等现象.各血清浓度培养体系中细胞生长曲线测定结果显示,在细胞培养的0、24、48、72、96 h,各组细胞密度与常规培养的对照组差别不明显.因此用MEM培养基培养IBRS-2细胞时,血清最低含量为2%.用该体系培养IBRS-2细胞接种PPV N株后,通过TCID50和HA测定病毒滴度.结果显示,2%血清培养的PPV N株病毒滴度平均值与常规条件培养(6%～10%血清)无明显差别,表明该体系可用于PPV N株的增殖.本研究成功驯化出能在低含量血清培养条件下生长良好的IBRS-2细胞,血清浓度可以降低到2%,且PPV N株在该低含量血清培养的IBRS-2细胞中能良好增殖.